This is a continuation of a grant that has concentrated on Streptococcus pneumoniae and its ability to adapt to the nasopharynx with colonization as well as to produce invasive infection with bacteremia. Such adaptation has been shown to be correlated with the expression of two distinct phenotypes, the opaque and transparent colony forms. Opacity phenotype is associated with on-off switching of pyruvate oxidase (SpxB), that mediates the aerobic metabolism of pyruvate and results in the generation of H2O2. Pursuing the biochemical and genetic basis of opacity variation will continue with a focus on the ability of the pneumococcus to produce and tolerate unusually high levels of H2O2. The overall goal of the proposal is to define the effect of opacity phenotype on carriage, the important first step in the pathogenesis of pneumococcal disease. The specific aims are: (1) to define the contribution of H2O2 production and opacity variation to pneumococcal carriage; (2) to characterize the physiology of H2O2 production; (3) to define the mechanism for resistance to high-level H2O2 production. A genetic approach here will be used to attempt to isolate sequences contributing to the resistance to endogenously generated peroxide; and (4) to identify and characterize regulatory elements controlling opacity (phenotypic) variation. The hope here is to isolate transcription factors that may function as global regulators affecting pyruvate oxidase.